摘要
目的探讨钙敏感受体(CaSR)过表达对缺血后未成熟脑白质祖细胞增殖分化的促进作用。方法从5 d龄SD大鼠脑室周围白质组织中提取、培养脑白质祖细胞,并分为对照组、氧糖剥夺(OGD)组、OGD+三氯化钆(GdCl3)组和OGD+CaSR沉默组,其中利用GdCl3特异性激动CaSR表达,利用基因沉默方式抑制CaSR表达。OGD后24 h、48 h、72 h、7 d和14 d,采用实时荧光定量逆转录PCR(RT-qPCR)检测各组细胞中CaSR mRNA水平,采用免疫荧光双标染色检测各组细胞的分化情况;OGD后48 h细胞球形成时,利用倒置显微镜检测细胞增殖情况。结果(1)CaSR mRNA表达情况:OGD后48 h、72 h、7 d,OGD组细胞内CaSR mRNA表达量均明显高于对照组,差异有统计学意义(P<0.05)。OGD后48 h、72 h、7 d及14 d,OGD+GdCl3组CaSR mRNA表达量均明显高于对照组及OGD组,OGD+CaSR沉默组CaSR mRNA表达量均明显低于对照组及OGD组,差异均有统计学意义(P<0.05)。(2)细胞增殖分化情况:OGD后48 h时,OGD组细胞球直径[(75.26±26.07)μm]较对照组[(57.96±18.92)μm]明显增大,OGD+GdCl3组细胞球直径[(91.92±21.82)μm]较对照组及OGD组明显增大,而OGD+CaSR沉默组细胞球直径[(24.09±8.34)μm]较对照组及OGD组明显减小,差异均有统计学意义(P<0.05)。OGD后48 h、72 h时,OGD组中O4+/CaSR+少突胶质细胞前体细胞(OPCs)数目明显高于对照组,OGD+GdCl3组中O4+/CaSR+OPCs数目明显高于对照组及OGD组,OGD+CaSR沉默组中O4+/CaSR+OPCs数目明显低于对照组及OGD组,差异均有统计学意义(P<0.05)。结论CaSR过表达可进一步促进脑白质祖细胞增殖及分化为OPCs。
Objective To explore the promoted role of calcium-sensing receptor(CaSR)overexpression in proliferation and differentiation of immature white matter progenitor cells after ischemia in vitro.Methods Periventricular white matter cells from 5-d-old rats were cultured in vitro and divided into control group,oxygen-glucose deprivation(OGD)group,OGD+gadolinium chloride(GdCl3)group,and OGD+CaSR silenced group.CaSR expression was agitated by GdCl3,and CaSR gene expression was inhibited by gene silencing.CaSR mRNA levels 24,48 and 72 h,and 7 and 14 d after OGD were detected by real-time fluorescence quantification-PCR(RT-qPCR);cell proliferation 48 h after OGD was detected by inverted microscope,and cell differentiation 24,48,and 72 h,and 7 and 14 d was detected by double immunofluorescence staining.Results(1)CaSR mRNA expressions:CaSR mRNA expressions in OGD group 48 and 72 h and 7 d after OGD were statistically higher than those in control group(P<0.05).Forty-eight and 72 h,and 7 and 14 d after OGD,CaSR mRNA expressions in OGD+GDCL3 group were statistically higher than those in control group and OGD group(P<0.05);the CaSR mRNA expressions in the OGD+CaSR silenced group were significantly lower than those in the control group(P<0.05).(2)Cell proliferation and differentiation:48 h after OGD,the cell sphere diameter of OGD group([75.26±26.07]μm)was significantly increased as compared with that of control group([57.96±18.92]μm,P<0.05);the cell sphere diameter of OGD+GdCl3 group([91.92±21.82]μm)was significantly increased as compared with that of control group and OGD group(P<0.05);and the cell sphere diameter of OGD+CaSR silenced group([24.09±8.34]μm)was significantly shorter than that of control group and OGD group(P<0.05).At 48 and 72 h after OGD,the number of O4+/CaSR+olidoendrocyte precursor cells(OPCs)in OGD group was significantly larger than that in control group,that in OGD+GdCl3 group was significantly larger than that in control group and OGD group,and that in OGD+CaSR silenced group was significantly smal
作者
毛凤霞
雷梦园
程欣茹
张菊
徐千雅
时赞扬
韩素鸽
张茜
Mao Fengxia;Lei Mengyuan;Cheng Xinru;Zhang Ju;Xu Qianya;Shi Zanyang;Han Suge;Zhang Qian(Department of Neonatology,First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China;Department of Medical Examination,First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China)
出处
《中华神经医学杂志》
CAS
CSCD
北大核心
2021年第4期325-330,共6页
Chinese Journal of Neuromedicine
基金
河南省医学科技攻关计划省部共建项目(SBGJ2018040)
河南省国际科技合作计划项目(172102410017)。
关键词
钙敏感受体
脑白质祖细胞
少突胶质细胞前体细胞
未成熟脑
脑室周围白质软化
Calcium-sensing receptor
White matter progenitor cell
Oligodendrocyte precursor cell
Immature brain
Periventricular leukomalacia
